Design, Development, and Characterization of Gallium (III) Salophen Metallotherapeutics Targeting Heme Sensing and Iron Acquisition Pathways in Pseudomonas aeruginosa

Abstract

The development of new antibiotics is outpaced by the rise in multi-drug resistant (MDR) bacteria, creating a global health problem. Pseudomonas aeruginosa, one such bacterium, is labeled as a “critical priority” pathogen by the WHO for its resistance to treatment and prevalence in hospital-acquired infections and immunocompromised patients where it is often life threatening. Adding to this problem, most new discoveries are derivatives of existing antibiotic classes rather than new strategies. Newer approaches targeting bacterial pathways critical to infection but not survival outside the host are expected to exert less selective pressure and slow resistance onset. One such strategy is interfering with bacterial iron uptake and utilization, as iron is a key micronutrient with several iron-regulated virulence traits used to counter iron-sequestering defense mechanisms of the host. P. aeruginosa can shift between the acquisition of labile iron stores and the more abundant heme-bound iron at various stages of infection, so inhibitors targeting these pathways must account for this adaptability. One such approach to targeting iron utilization in several forms is the use of gallium, which mimics ferric iron in ionic size and charge but cannot undergo critical redox processes, thus causing toxicity in the bacteria that acquire it under the guise of iron. This work describes the synthesis and characterization of Gallium Salophen (GaSal) and subsequent analogs targeting heme and iron acquisition pathways in P. aeruginosa. In this characterization, GaSal binds to a hemophore, HasAp, secreted by P. aeruginosa, and inhibits an extra-cytoplasmic function (ECF) signaling cascade with the outer-membrane receptor HasR, which is critical for sensing and adapting to host heme levels. GaSal is simultaneously a substrate for uptake, independent of its effect on HasAp. Using a combination of cell-based assays as well as in vitro target characterization and finally preliminary animal infection studies, GaSal and subsequent derivatives are shown to be promising new developments targeting several points in the iron uptake and utilization pathways of P. aeruginosa. Continued developments aim to retain such activity and include several routes towards further optimization and development as a therapeutic.